Emerging human protoparvoviruses : in search of acute infections

Parvovirukset ovat eräitä pienimpiä tunnettuja viruksia. Parvovirusten genomi on yksijuosteinen DNA, jonka koko on noin viisituhatta emäsparia. Parvovirus virionilla on pieni (20-30 nm), vaipaton ikosahedraalinen kapsidi. Parvovirukset voivat aiheuttaa monenlaisia infektioita. Kenties tunnetuin ihmiselle patogeeninen parvovirus on parvovirus B19 (B19V), joka aiheuttaa parvorokkoa, anemioita ja sikiökuolemia. Toinen paljon tutkittu parvovirus on ihmisen bocavirus 1 (HBoV), joka aiheuttaa pienten lasten hengitystieinfektioita. Bufavirus (BuV), tusavirus (TuV) ja cutavirus (CuV) ovat uusia parvoviruksia, jotka löydettiin vuosina 2012–2016 syväsekvensointimenetelmillä. BuV löydettiin alun perin ulostenäytteestä Burkina Fasosta ja virusta on sen jälkeen löydetty polymeraasiketjureaktioon (PCR) perustuvien menetelmien avulla ulostenäytteistä Euroopasta, Afrikasta ja Aasiasta. BuV:n seroprevalenssi vaihtelee alueittain. TuV löydettiin yhdestä ainoasta ulostenäytteestä Tunisiasta, eikä TuV DNA:ta ole löydetty sen jälkeen muista näytteistä. CuV löydettiin vuonna 2016 ja se on yhdistetty kutaaniseen T-solulymfoomaan, mutta ei tiedetä aiheuttaako virus syövän vai hakeutuuko se vain nopeasti jakautuviin syöpäsoluihin. BuV, TuV ja CuV kuuluvat Protoparvovirus sukuun, mutta ei tiedetä onko TuV ihmisen virus. Kaikkien kolmen uuden parvoviruksen tutkimus on vielä alussa. On paljon, mitä ei tiedetä näiden kolmen viruksen epidemiologiasta ja niiden aiheuttamista oireista. Tämän pro gradu – työn kahtena päätavoitteena oli: pystyttää µ-kaappaus-IgM-entsyymivälitteinen immunosorbenttimääritys (EIA) -testi ihmisen protoparvoviruksille käyttäen BuV1:ta esimerkkinä sekä analysoida kolme ulostenäytekohorttia valmiilla BuV, TuV ja CuV kvantitatiivisella multiplex PCR (qPCR)-menetelmällä. IgM EIA:n pystytyksen perustana käytettiin aiemmin HBoV:lle ja parvovirus B19V:lle pystytettyjä µ-kaappaus-IgM-EIA-testejä. Menetelmää jouduttiin optimoimaan paljon ja lopulta monia testauksia jouduttiin tekemään, sillä menetelmä ei toiminut kuten oli oletettu. Vian etsinnän tuloksena saatiin selville, että epäselvät tulokset saattoi aiheuttaa hajonneet VLP:t tai, että herkkä µ-kaappaus-testi vaatii erityisen puhtaita VLP:tä. Lopullista toimivaa µ-kaappaus-IgM-EIA-menetelmää protoparvoviruksille ei tässä työssä pystytty pystyttämään, mutta tämä työ luo hyvän pohjan jatkokehitykselle. BuV, TuV ja CuV DNA:ta löydettiin ulostenäytteistä multiplex qPCR-menetelmällä. QPCR- ja sekvensointitulosten perusteella tässä työssä löydettiin yksi BuV DNA-positiivinen ja yksi TuV DNA positiivinen näyte. Lisäksi löydettiin 12 CuV DNA positiivista näytettä. TuV DNA:ta havaittiin ensi kertaa sen ensilöydön jälkeen. Sen lisäksi, CuV DNA:ta löydettiin ensilöydön jälkeen jälleen ulostenäytteestä. Useissa aiemmissa tutkimuksissa CuV DNA:ta on löydetty ihosta otetuista biopsioista. Fylogeneettisten analyysien perusteella TuV:n sekvenssi on lähempänä jyrsijöiden parvoviruksia kuin BuV:n ja CuV:n sekvenssit. Tutkimusta ja erilaisia näytteitä (eläin- ja ihmisperäisiä) tarvitaan, jotta voidaan selvittää onko TuV ihmisen virus.Parvoviruses are among the smallest known viruses. The parvovirus genome is a single stranded DNA, approximately 5 kb in size. The virion has a small (20 to 30 nm), rugged, non-enveloped icosahedral capsid. Parvoviruses can cause a number of diseases. Possibly the most recognized human parvovirus is parvovirus B19 (B19V), which can cause the so-called fifth disease, anemias and fetal death. Another relatively well characterised parvovirus is human bocavirus 1 (HBoV1), which causes respiratory tract infections in young children. Bufavirus (BuV) tusavirus (TuV) and cutavirus (CuV) are emerging parvoviruses, discovered during the years 2012-2016 using next generation sequencing methods. All three viruses were originally discovered in feces of patients suffering from diarrhea. BuV was originally found in Burkina Faso and has since been detected in fecal samples with polymerase chain reaction (PCR)-based methods from Europe, Asia and Africa. The seroprevalence of BuV differs between countries. TuV was found in a single stool sample from Tunisia, but no further reports of it have since emerged. CuV was found in 2016 and it has been linked to cutaneous T-cell lymphoma, but it is not known if the virus is the cause of the cancer or if the virus simply prefers quickly dividing cancer cells for its replication. BuV, TuV and CuV belong to the Protoparvovirus genus, but it is still unclear whether TuV is a human pathogen. More research is needed to study the epidemiology of these viruses and their role in illnesses. There were two main aims in this thesis: to set up an IgM µ-capture enzyme immunoassay (EIA) for human protoparvoviruses using BuV1 as an example and to screen three stool sample cohorts for BuV, TuV and CuV using an in-house multiplex quantitative PCR (qPCR). The IgM EIAs developed for B19V and HBoV1 was used as the base for developing human protoparvovirus IgM EIA, using Virus-like particles (VLP) as antigens. Setting up the EIA required a great amount of optimization and finally troubleshooting, since the assay did not work as expected. The troubleshooting revealed that the ambiguous results in the IgM µ-capture EIA were possibly due to degraded VLPs or that the sensitive µ-capture format requires extremely carefully purified VLPs. More optimizing is needed for this assay, however, the work done in this thesis offers a good base for further development of protoparvovirus IgM EIA. All three viruses were found in the stool samples during multiplex qPCR screening. Based on the qPCR and sequencing results one sample was positive for BuV DNA, one sample for TuV DNA and a total of 12 samples for CuV DNA. This is the first time TuV DNA has been found since its discovery. In addition to that, CuV DNA was identified in fecal samples for the first time since the discovery, previously CuV DNA had been found mostly in skin biopsies. As for TuV, based on the parvovirus phylogenetic analyses, its sequence is more closely related to rodent parvoviruses than CuV or BuV. More research is needed, possibly with animal and human samples, to establish the role of TuV as a human virus

Kaupunkiluonto ja monikulttuurisuus – maahanmuuttajat luontoalueiden kokijoina ja käyttäjinä

Kaupungistuminen ja maahanmuutosta seuraava monikulttuuristuminen ovat merkittäviä yhteiskunnallisia muutoksia. Kaupunkiluonnon merkitys ihmisten hyvinvoinnille korostuu väestön keskittyessä kasvaville kaupunkiseuduille, mutta kaupunkiluonnon merkitystä maahanmuuttajien integroitumisessa ei ole juurikaan tutkittu. Tässä raportissa: - Kartoitetaan, miten eri maahanmuuttajaryhmien määrä on kehittymässä Helsingin seudulla ja mitkä ovat suuria tai kasvavia ja siten erityisesti huomionarvoisia ryhmiä. - Selvitetään, mitä tietoa on saatavilla ihmisten luontosuhteen vaihtelusta eri kulttuureissa. - Selvitetään, millainen merkitys kaupunkiluonnolla on maahanmuuttajien integroitumisessa uuteen elinympäristöön ja yhteisöön. - Muodostetaan käsitys keskeisistä tiedon puutteista ja tutkimustarpeista. Maahanmuuttajien määrän kehitystä Helsingin seudulla tarkastellaan tilastojen avulla. Lisäksi arvioidaan tilastojen käyttöön liittyviä haasteita. Eri kulttuuritaustaisten ryhmien luontoon liittämiä merkityksiä ja luonnon käytön tapoja tarkastellaan aiempien tutkimusten perusteella. Maahanmuuttajien luontosuhteeseen keskittyvää kotimaista tutkimusta ei juuri löytynyt, mutta aiheeseen liittyvistä tutkimuksista käy ilmi arvojen ja kokemusten vaihtelu kulttuurien välillä. Julkaisussa esitellään esimerkkitapauksia luonnon merkityksistä maahanmuuttajien integroitumisessa ja pohditaan integroitumisen edistämisen mahdollisuuksia Helsingin seudulla. Pyrittäessä edistämään yksilöiden ja ryhmien integroitumista on tärkeää tunnistaa luontoalueiden mahdollisuuksia symbolisen tunnistamisen ja sosiaalisen vuorovaikutuksen luomisessa. Tietoa tarvitaan myös siitä, millä tavoin näitä mahdollisuuksia voidaan tukea luontoalueiden maankäytön ja hoidon suunnittelun kautta. Maahanmuuttajien näkemysten ja kokemusten hyödyntäminen voi auttaa kaupunkien suunnittelua löytämään muuttuviin olosuhteisiin sopivia toimintatapoja ja ratkaisuja

Exploring the association between canine perineal hernia and neurological, orthopedic, and gastrointestinal diseases

Publisher Copyright: © 2022, The Author(s).Background: Perineal hernia (PH) is a relatively common condition in intact male dogs, but the etiology remains unclear. The objective of this study was to assess the contribution of gastrointestinal (GI), neurological, and orthopedic conditions to the development of PH in male dogs. Patient history with a focus on chronic GI disease was assessed using an owner questionnaire. Neurological conditions were explored, applying neurological, electromyographic (EMG), and motor nerve conduction velocity (MNCV) examinations and combining these with computed tomography (CT) imaging. To exclude possible orthopedic diseases, an orthopedic examination was conducted together with CT analysis. The chi-squared test was used to assess the associations between categorical variables. Results: Altogether, 66 male dogs with diagnosed PH were recruited for this study. The frequency of neurological, orthopedic, and GI diseases was low in dogs with PH. No signs of generalized neuro- or myopathies were detected. Still, perineal and bulbourethral reflexes were decreased or missing in 44.6% (29/65) and 40.0% (26/65) of dogs, respectively. Mild or moderate occlusion of the intervertebral foramen at the lumbosacral (LS) junction occurred in 18.5% (12/65) of dogs and was caused by spondylosis deformans in 83.3% (10/12). Moderate disc protrusion was evident in 9.2% (6/65) of dogs. Conclusion: No evidence was found that PH is caused by gastrointestinal, orthopedic, or neurological conditions. Abnormalities in perineal and bulbourethral reflexes are most likely secondary to PH.Peer reviewe

Human Protoparvovirus DNA and IgG in Children and Adults with and without Respiratory or Gastrointestinal Infections

Abstract: Three human protoparvoviruses, bufavirus (BuV), tusavirus (TuV) and cutavirus (CuV), have recently been discovered in diarrheal stool. BuV has been associated with diarrhea and CuV with cutaneous T-cell lymphoma, but there are hardly any data for TuV or CuV in stool or respiratory samples. Hence, using qPCR and IgG enzyme immunoassays, we analyzed 1072 stool, 316 respiratory and 445 serum or plasma samples from 1098 patients with and without gastroenteritis (GE) or respiratory-tract infections (RTI) from Finland, Latvia and Malawi. The overall CuV-DNA prevalences in stool samples ranged between 0–6.1% among our six patient cohorts. In Finland, CuV DNA was significantly more prevalent in GE patients above rather than below 60 years of age (5.1% vs 0.2%). CuV DNA was more prevalent in stools among Latvian and Malawian children compared with Finnish children. In 10/11 CuV DNA-positive adults and 4/6 CuV DNA-positive children with GE, no known causal pathogens were detected. Interestingly, for the first time, CuV DNA was observed in two nasopharyngeal aspirates from children with RTI and the rare TuV in diarrheal stools of two adults. Our results provide new insights on the occurrence of human protoparvoviruses in GE and RTI in different countries.Peer reviewe

Human Protoparvovirus DNA and IgG in Children and Adults with and without Respiratory or Gastrointestinal Infections

Abstract: Three human protoparvoviruses, bufavirus (BuV), tusavirus (TuV) and cutavirus (CuV), have recently been discovered in diarrheal stool. BuV has been associated with diarrhea and CuV with cutaneous T-cell lymphoma, but there are hardly any data for TuV or CuV in stool or respiratory samples. Hence, using qPCR and IgG enzyme immunoassays, we analyzed 1072 stool, 316 respiratory and 445 serum or plasma samples from 1098 patients with and without gastroenteritis (GE) or respiratory-tract infections (RTI) from Finland, Latvia and Malawi. The overall CuV-DNA prevalences in stool samples ranged between 0–6.1% among our six patient cohorts. In Finland, CuV DNA was significantly more prevalent in GE patients above rather than below 60 years of age (5.1% vs 0.2%). CuV DNA was more prevalent in stools among Latvian and Malawian children compared with Finnish children. In 10/11 CuV DNA-positive adults and 4/6 CuV DNA-positive children with GE, no known causal pathogens were detected. Interestingly, for the first time, CuV DNA was observed in two nasopharyngeal aspirates from children with RTI and the rare TuV in diarrheal stools of two adults. Our results provide new insights on the occurrence of human protoparvoviruses in GE and RTI in different countries.Peer reviewe

HERQ-9 Is a New Multiplex PCR for Differentiation and Quantification of All Nine Human Herpesviruses

Infections with the nine human herpesviruses (HHVs) are globally prevalent and characterized by lifelong persistence. Reactivations can potentially manifest as life-threatening conditions for which the demonstration of viral DNA is essential. In the present study, we developed HERQ-9, a pan-HHV quantitative PCR designed in triplex reactions to differentiate and quantify each of the HHV-DNAs: (i) herpes simplex viruses 1 and 2 and varicella-zoster virus; (ii) Epstein-Barr virus, human cytomegalovirus, and Kaposi's sarcoma-associated herpesvirus; and (iii) HHV-6A, -6B, and -7. The method was validated with prequantified reference standards as well as with mucocutaneous swabs and cerebrospinal fluid, plasma, and tonsillar tissue samples. Our findings highlight the value of multiplexing in the diagnosis of many unsuspected, yet clinically relevant, herpesviruses. In addition, we report here frequent HHV-DNA co-occurrences in clinical samples, including some previously unknown. HERQ-9 exhibited high specificity and sensitivity (LOD95 of similar to 10 to similar to 17 copies/reaction), with a dynamic range of 10' to 10 6 copies/p.I. Moreover, it performed accurately in the coamplification of both high- and low-abundance targets in the same reaction. In conclusion, we demonstrated that HERQ-9 is suitable for the diagnosis of a plethora of herpesvirus-related diseases. Besides its significance to clinical management, the method is valuable for the assessment of hitherto-unexplored synergistic effects of herpesvirus coinfections. Furthermore, its high sensitivity enables studies on the human virome, often dealing with minute quantities of persisting HHVs. IMPORTANCE By adulthood, almost all humans become infected by at least one herpesvirus (HHV). The maladies inflicted by these microbes extend beyond the initial infection, as they remain inside our cells for life and can reactivate, causing severe diseases. The diagnosis of active infection by these ubiquitous pathogens includes the detection of DNA with sensitive and specific assays. We developed the first quantitative PCR assay (HERQ-9) designed to identify and quantify each of the nine human herpesviruses. The simultaneous detection of HHVs in the same sample is important since they may act together to induce life-threatening conditions. Moreover, the high sensitivity of our method is of extreme value for assessment of the effects of these viruses persisting in our body and their long-term consequences on our health.Peer reviewe

Human Protoparvovirus DNA and IgG in Children and Adults with and without Respiratory or Gastrointestinal Infections

Three human protoparvoviruses, bufavirus (BuV), tusavirus (TuV) and cutavirus (CuV), have recently been discovered in diarrheal stool. BuV has been associated with diarrhea and CuV with cutaneous T-cell lymphoma, but there are hardly any data for TuV or CuV in stool or respiratory samples. Hence, using qPCR and IgG enzyme immunoassays, we analyzed 1072 stool, 316 respiratory and 445 serum or plasma samples from 1098 patients with and without gastroenteritis (GE) or respiratory-tract infections (RTI) from Finland, Latvia and Malawi. The overall CuV-DNA prevalences in stool samples ranged between 0-6.1% among our six patient cohorts. In Finland, CuV DNA was significantly more prevalent in GE patients above rather than below 60 years of age (5.1% vs 0.2%). CuV DNA was more prevalent in stools among Latvian and Malawian children compared with Finnish children. In 10/11 CuV DNA-positive adults and 4/6 CuV DNA-positive children with GE, no known causal pathogens were detected. Interestingly, for the first time, CuV DNA was observed in two nasopharyngeal aspirates from children with RTI and the rare TuV in diarrheal stools of two adults. Our results provide new insights on the occurrence of human protoparvoviruses in GE and RTI in different countries

HERQ-9 Is a New Multiplex PCR for Differentiation and Quantification of All Nine Human Herpesviruses

Infections with the nine human herpesviruses (HHVs) are globally prevalent and characterized by lifelong persistence. Reactivations can potentially manifest as life-threatening conditions for which the demonstration of viral DNA is essential. In the present study, we developed HERQ-9, a pan-HHV quantitative PCR designed in triplex reactions to differentiate and quantify each of the HHV-DNAs: (i) herpes simplex viruses 1 and 2 and varicella-zoster virus; (ii) Epstein-Barr virus, human cytomegalovirus, and Kaposi’s sarcoma-associated herpesvirus; and (iii) HHV-6A, -6B, and -7. The method was validated with prequantified reference standards as well as with mucocutaneous swabs and cerebrospinal fluid, plasma, and tonsillar tissue samples. Our findings highlight the value of multiplexing in the diagnosis of many unsuspected, yet clinically relevant, herpesviruses. In addition, we report here frequent HHV-DNA co-occurrences in clinical samples, including some previously unknown. HERQ-9 exhibited high specificity and sensitivity (LOD95s of ∼10 to ∼17 copies/reaction), with a dynamic range of 101 to 106 copies/μl. Moreover, it performed accurately in the coamplification of both high- and low-abundance targets in the same reaction. In conclusion, we demonstrated that HERQ-9 is suitable for the diagnosis of a plethora of herpesvirus-related diseases. Besides its significance to clinical management, the method is valuable for the assessment of hitherto-unexplored synergistic effects of herpesvirus coinfections. Furthermore, its high sensitivity enables studies on the human virome, often dealing with minute quantities of persisting HHVs.IMPORTANCE By adulthood, almost all humans become infected by at least one herpesvirus (HHV). The maladies inflicted by these microbes extend beyond the initial infection, as they remain inside our cells for life and can reactivate, causing severe diseases. The diagnosis of active infection by these ubiquitous pathogens includes the detection of DNA with sensitive and specific assays. We developed the first quantitative PCR assay (HERQ-9) designed to identify and quantify each of the nine human herpesviruses. The simultaneous detection of HHVs in the same sample is important since they may act together to induce life-threatening conditions. Moreover, the high sensitivity of our method is of extreme value for assessment of the effects of these viruses persisting in our body and their long-term consequences on our health.</p

Differential Mitochondrial Gene Expression in Adipose Tissue Following Weight Loss Induced by Diet or Bariatric Surgery

Context: Mitochondria are essential for cellular energy homeostasis, yet their role in subcutaneous adipose tissue (SAT) during different types of weight-loss interventions remains unknown. Objective: To investigate how SAT mitochondria change following diet-induced and bariatric surgery-induced weight-loss interventions in 4 independent weight-loss studies. Methods: The DiOGenes study is a European multicenter dietary intervention with an 8-week low caloric diet (LCD; 800 kcal/d; n = 261) and 6-month weight-maintenance (n = 121) period. The Kuopio Obesity Surgery study (KOBS) is a Roux-en-Y gastric bypass (RYGB) surgery study (n = 172) with a 1-year follow-up. We associated weight-loss percentage with global and 2210 mitochondria-related RNA transcripts in linear regression analysis adjusted for age and sex. We repeated these analyses in 2 studies. The Finnish CRYO study has a 6-week LCD (800-1000 kcal/d; n = 19) and a 10.5-month follow-up. The Swedish DEOSH study is a RYGB surgery study with a 2-year (n = 49) and 5-year (n = 37) follow-up. Results: Diet-induced weight loss led to a significant transcriptional downregulation of oxidative phosphorylation (DiOGenes; ingenuity pathway analysis [IPA] z-scores: -8.7 following LCD, -4.4 following weight maintenance; CRYO: IPA z-score: -5.6, all P < 0.001), while upregulation followed surgery-induced weight loss (KOBS: IPA z-score: 1.8, P < 0.001; in DEOSH: IPA z-scores: 4.0 following 2 years, 0.0 following 5 years). We confirmed an upregulated oxidative phosphorylation at the proteomics level following surgery (IPA z-score: 3.2, P < 0.001). Conclusions: Differentially regulated SAT mitochondria-related gene expressions suggest qualitative alterations between weight-loss interventions, providing insights into the potential molecular mechanistic targets for weight-loss success.Peer reviewe